Copines are highly conserved calcium-dependent membrane-binding proteins found in numerous diverse eukaryotic organisms, including humans. Studies with human copines suggest that copines may act in signaling pathways by binding to target proteins and then recruiting these target proteins to a particular membrane in response to an increase in calcium concentration. This proposal describes experiments to specifically test the above hypothesis using developing Dictyostelium as a model. Dictyostelium can live independently as single celled amoebae, but when placed in starvation conditions, single amoeba signal each other to first aggregate and then differentiate into cells that form a multicellular fruiting body. This simple developmental program in Dictyostelium is widely used to study not only development, but also several basic cell processes including chemotaxis-mediated cell motility, signal transduction, programmed cell death cell, and cell differentiation. Previous studies show that one of the copine proteins in Dictyostelium, CpnA, is required for normal development and the developmental expression patterns of several of the other copine genes suggest they too may be regulators of development. The experiments outlined in specific aim 1 are designed to further characterize the role of CpnA in development as well as investigate the function of two other copine proteins. These experiments will allow us to determine the developmental and cellular processes in which copines function; however, the exact mechanism that they use to carry these functions will require biochemical studies like the ones proposed in the second specific aim. Experiments outlined in specific aim 2 are designed to identify the target proteins of the Dictyostelium copines to explore the mechanistic role of copines as chaperone or recruiting proteins functioning in signaling pathways. The human genome has eight copine genes and most of them are expressed in bone marrow cells and blood leukocytes; therefore, copines are potentially involved in cell signaling pathways within these cells. Increasing our scientific knowledge of how copines function could lead to a better understanding of human blood and immune cell-related diseases. PUBLIC HEALTH RELEVANCE: The proposal outlines experiments to investigate the functions of a novel family of proteins called copines. Copines are hypothesized to have roles in cell signaling pathways, but their exact functions are unknown. The human genome has eight copine genes and most are found in bone marrow cells and blood leukocytes. Increasing our scientific knowledge of how copines function could potentially lead to a better understanding of human blood and immune cell related diseases. [unreadable] [unreadable] [unreadable] [unreadable]